The present invention relates to compositions, methods and devices for the detection of infection by endogenous retroviruses. In particular, the present invention comprises compositions, devices and methods for the detection of porcine endogenous retroviruses, detection of which is necessary following xenotransplantation of porcine cellular products.
Currently, there are shortages of human organs and cells for transplantation into humans. These shortages of human donor material suitable for allotransplantation, coupled with recent advances in transplantation immunology, have provided impetus for attempts to develop xenotransplantationxe2x80x94the therapeutic use of living animal tissues and organs in humans.
To date, the therapeutic promise of xenotransplantation has not become widely accepted. There have been incidences of transplantation of simian organs and porcine cells and organs. Progress has been encouraging enough to merit the beginning of limited clinical trials in the United States.
Pigs are among the primary animal species proposed as sources of xenografts. Xenotransplantation clinical trials involving porcine tissue being considered or underway include, the perfusion through or implantation of whole liver preparations as a treatment for hepatic failure, the implantation of fetal neuronal tissue as a therapy for Parkinson""s disease, and the infusion or implantation of pancreatic islet cells as a treatment for diabetes mellitus. Concerns have been raised that the implantation of porcine tissue and/or cells into immune compromised humans may facilitate the transmission of new infectious agents to humans. The Public Health Service has therefore stressed the importance of proceeding with xenotransplantation clinical trials only after there are diagnostic tools that detect infectious agents from pigs, surveillance programs for new xenograft recipients have been developed, and persons exposed to xenografts can be tested for evidence of infection.
There is such a need for detection and monitoring because porcine tissues and cells are infected with endogenous retroviruses. The genomes of all domesticated swine species tested thus far contain multiple integrated copies of an endogenous C-type retrovirus termed porcine endogenous retrovirus (PERV). Recently PERV has been reported to be transmissible to human cell lines in vitro, thus raising concerns that PERV may be capable of infecting recipients of porcine xenografts. The potential infection of human transplant recipients with new xenogeneic infectious agents, and subsequent transmission of these infections to the general population is a major concern when using animal tissue for human transplantation. Risks for xenogeneic infections may be significantly increased by the immunosuppressive therapies required to maintain graft function in human xenotransplant recipients.
One possible approach to detection of these xenogeneic infectious agents, including endogenous retroviruses, is to use molecular biological techniques. Recent sequence analysis of PERV genomes has allowed the development of diagnostic polymerase chain reactions (pcr) assays to monitor for the presence of virus in peripheral blood lymphocytes, sera or other tissues. However, these assays are limited to detecting infection when virus is present in the patient material sampled. An integrated copy of the PERV genome, that can be activated later within the human transplant recipient, cannot be detected except with PCR methods and then only if virus is present. Neither the potential for PERV to infect humans nor tissue tropism of PERV have been determined, and thus, it is not certain if human infection would be easily detectable by molecular approaches.
Prior to the present invention, there has been no development of a serologic assay that is capable of detecting PERV. The development of a specific serology assay has been problematic, due in part to a lack of an appropriate positive control antisera. Lack of antisera that only identifies PERV antigens, and not porcine or other cellular products has hampered advances in serologic detection methods.
Although immune sera from an animal infected with PERV would be ideal, cross-species infections by PERV have not yet been found. Pigs are immunologically tolerant to PERV and thus, do not make antibodies that are specific for viral antigens. Raising antisera by immunization with whole virus lysate would be a possible approach, however until very recently the only potential source of PERV antigens have been porcine cell lines constitutively expressing the virus. The preparation of viral proteins, free of contaminating porcine antigens, would be extremely difficult if not impossible to achieve from porcine-derived cells.
One method for stopping the transmission of porcine endogenous viruses from xenotransplantation would be to harvest the transplantation materials from virus-free animals. The risks of transmission of known infectious agents may be reduced, or eliminated by procuring source animals from specific pathogen-free colonies. However, this pre-transplant screening method cannot eliminate the porcine endogenous retrovirus (PERV), because the genome of these viruses is carried in the germ line of every pig. Pig PERV particles of type C morphology are released spontaneously by cell lines originating from a variety of pig tissues including kidneys, lymph nodes, testes and fallopian tubes. All known PERVs originate from healthy porcine tissues except for two known. types, PERV-Shimozuma-1 and 38A-1 which are derived from porcine lymphomas.
The knowledge that PERV originating from both cell lines and primary porcine lymphocytes can infect human cells in vitro has heightened safety concerns related to pig xenografts. Transmission of xenogeneic retroviral infections to xenograft recipients is of particular concern because retroviruses are known to result in life long persistent infections. The current absence of the ability to detect the presence of PERV, which hampers the determination of whether PERV will infect humans exposed to porcine xenografts, and whether PERV will be transmitted secondarily among their contacts, has raised questions on the safety of pig-to-human transplantation, and threatens to delay progress in this therapeutic technology.
Thus, there is a long felt need for methods, devices and compositions that are capable of detecting the presence of endogenous viruses in transplantation materials or that may be released from implanted tissues. Particularly needed, are compositions and methods for diagnostic and monitoring assays that are capable of detecting the presence of antigens of endogenous viruses, particularly PERV antigens. Such assays would be important in providing vital diagnostic and physiologic information for xenotransplant recipients.
In accordance with the present invention, compositions, methods and devices are provided that are effective in serologic detection of endogenous retroviruses. These methods and compositions are particularly effective in detecting type-C retroviruses, and more particularly are effective in detecting porcine endogenous retrovirus, PERV. These compositions and methods can be used in diagnostic devices for monitoring and detecting the presence of porcine endogenous retrovirus (PERV) in transplant recipients.
Accordingly, it is an object of the present invention to provide compositions that are capable of detecting type-C retroviruses.
It is yet another object of the present invention to provide methods and compositions for detecting type-C retroviruses.
Another object of the present invention to provide methods and compositions for serologic assays for detection of type-C retroviruses.
A further object of the present invention to provide compositions that are capable of detecting porcine endogenous retrovirus (PERV).
It is yet another object of the present invention to provide methods for detecting porcine endogenous retrovirus (PERV).
Another object of the present invention to provide serologic assay devices for detection of porcine endogenous retrovirus (PERV).
It is yet another object of the present invention to provide compositions, methods and devices for monitoring xenotransplant recipients for the presence of xenogeneic endogenous retroviruses.
It is an object of the present invention to provide compositions, methods and devices for detecting the presence of endogenous retroviruses in transplant material, including organs, tissues and cells.
Still another object of the present invention is to provide compositions, methods and devices for determining the zoonotic potential and tissue tropism of endogenous retroviruses.
These and other objects, features and advantages of the present invention will become apparent after a review of the following detailed description of the disclosed embodiments and the appended claims.